Mutations of the type IV collagen alpha 1 gene (Col4a1) can cause anterior segment dysgenesis (ASD) and elevated intraocular pressure in mice. Recently, a COL4A1 mutation was identified in a family with ASD, ocular hypertension and juvenile glaucoma, however, COL4A1, has not widely been considered a candidate gene for ASD. We hypothesize that mutations of COL4A1 and its binding partner COL4A2 underlie ASD in patients for whom mutations have not yet been identified. In Aim 1, we will test our hypothesis directly by performing mutational analysis on ASD patients that are known not to have mutations in other ASD-causing genes. Moreover, we seek to determine where, and by what cellular mechanism(s), Col4a1 mutation leads to ASD. We have determined that COL4A1 is ubiquitously present in ocular basement membranes during development, which complicates the task of determining sites of primary pathogenesis from secondary pathology. To overcome this obstacle we have developed and validated a conditional mutant allele of Col4a1. In Aim 2, we will express the mutation only in the developing lens or only in cells of neural crest origin to determine their relative contributions to ASD. Finally, to understand how Col4a1 mutations lead to ocular dysgenesis and to begin to assemble developmental and pathogenic pathways, we will identify genetic modifiers of ASD. We have already successfully mapped a locus that is able to strongly rescue ASD and permit mutant mice to have nearly normally developed eyes. In Aim 3, we will perform systematic large-scale screens of two distinct genetic backgrounds to identify additional dominant or recessive modifier loci and genes. Understanding how genetic modifiers rescue disease could provide valuable insight for how targeted therapeutic interventions might do the same. To our knowledge, no other group is currently evaluating the role of COL4A1 or COL4A2 in ocular development and disease. The experiments outlined in this proposal take advantage of valuable and unique resources and will provide important new insights into the mechanisms of normal ocular development and pathogenic pathways. PUBLIC HEALTH RELEVANCE: This application seeks to identify a novel genetic cause of ocular dysgenesis that leads to early onset and aggressive glaucoma. The study will use unique genetic tools to understand how these genes lead to disease and which cellular pathways might be targeted with novel therapeutic interventions.